Development of an effective intervention strategy against congenital cytomegalovirus (CMV) is a major public health priority. However, candidate CMV vaccines strategies have been unsuccessful in completely preventing congenital infection in a pre-clinical animal model and an effective vaccine remains an elusive goal. The pathogenicity of CMV transplacental infection is poorly defined and a better understanding may provide fresh insight for the development of a novel intervention or vaccine strategy. In this proposal, we will examine two key aspects of placental infection: (1) Toll-like receptor (TLR) mediated innate immune response of the placenta to CMV; (2) Viral tropism and placental infection. The inflammatory innate immune response mediated via TLRs expressed on key placental cells (trophoblasts) is potentially an important aspect of controlling placental invasion congenital infection by both bacteria and viruses. CMV is known to be recognized by multiple TLRs but the placental TLR mediated response to CMV is poorly defined as is the mechanism by which CMV potentially circumvents the placental innate immunity and infects the fetus in utero. It is our hypothesis that successful congenital infection is reliant upon the ability of CMV to efficiently enter and usurp TLR expressing trophoblast cells, an important component of the placental barrier and placental innate immunity. Potentially, the newly identified endocytic pathway of viral cell entry could be an important factor in the invasion of the placenta and infection of epithelial trophoblasts. We propose to determine the significance of the TLR mediated innate immune response of the placenta to CMV infection in the only small animal model (guinea pig) for congenital CMV. Additionally, using the same animal model, we will define important tropism genes that potentially enable the virus to establish infectious foci in th placenta and subsequently infect the fetus in utero. These studies will employ conventional histopathological and immunohistochemical approaches as well as novel techniques including the use of bioluminescence imaging of viral dissemination in the animal model.